Pheromones and pheromone receptors have long been known to affect virulence of the opportunistic pathogen Cryptococcus neoformans, yet it is still entirely unclear how the biological ramifications of pheromone signaling affect pathogenesis. The Nielsen laboratory has shown that production of a newly characterized cell type - referred to as titan cells - is altered by pheromone sensing. Over-production of titan cells is correlated with reduced CNS penetration. The objective of this application is to define the role of titan cells in pathogenesis and how pheromone signaling alters titan cell production. The central hypothesis of our application is that the pheromone signaling cascade, in conjunction with other signaling components, regulates titan cell formation, which affects subsequent dissemination to the CNS. The rationale for the proposed research is that, once the role of titan cells in pathogenesis and how they are regulated is known, titan cell production can be targeted in new and innovative approaches for the prevention and treatment of cryptococcosis. We plan to test our central hypothesis and accomplish the objective of this application by pursuing the following specific aims: 1) Establish the role of titan cells in pathogenesis, and 2) Identify signaling pathways involved in titan cell formation. The expected outcomes of this research are to answer the long-standing question of why pheromone signaling affects pathogenesis in C. neoformans, to define the effect of titan cell production on pathogenesis, to identify signals that lead to titan cell production, and to understand titan cell formation and replication. Such results will have an important positive impact, because identified signaling components and/or downstream targets important for titan cell formation are expected to provide new targets for preventative and therapeutic interventions that will aid HIV/AIDS patients. We expect our contribution to be a detailed understanding of how signal transduction cascades regulate titan cell formation and how titan cell formation affects subsequent cell dissemination to the CNS. The proposed research is significant because it is expected to provide knowledge about titan cell formation, regulation, and pathogenesis that will enhance our understanding of the fundamental requirements for cryptococcal disease and allow titan cells to be targeted by antifungal therapy.